一般演題（口述）

一般演題（口述） Synapse

3O1-01 Molecular mechanism of monoamine deficiency in the mouse lacking an enzyme for recycling of tetrahydrobiopterin Ichinose Hiroshi，Xu Feng，Sudo Yusuke，Sanechika Sho，Hara Yoshitaka Grad School of Bioscience and Biotechnology, Tokyo Institute of Technology Quinonoid dihydropteridine reductase（QDPR）regenerates tetrahydrobiopterin（BH4）, which is a cofactor for monoamine synthesis and phenylalanine metabolism. Patients who have a genetic mutation in the QDPR gene develop hyperphenylalaninemia and severe neurological symptoms including dystonia, convulsion, and hyperthermia due to the depletion of catecholamines and serotonin. We examined a Qdpr-deficient mouse model in order to reveal the physiological significance of the BH4 recycling reaction. The Qdpr-deficient mice showed mild hyperphenylalaninemia and monoamine deficiency, although the BH4 contents in the liver and brains were not decreased. The blood phenylalanine levels were dropped off by the intraperitoneal injection of BH4, indicating the lack of BH4 in the liver. The serotonin contents in the brain were slightly increased after the administration of BH4, whereas the dopamine and noradrenaline contents in the brain were unchanged. Then, we treated the Qdpr-deficient mice with a phenylalanine-restricted diet. The brain monoamine levels were restored by the diet. Because the high concentration of phenylalanine can competitively inhibit transportation of tyrosine and tryptophan into the brain through L-type amino acid transporter 1, and inhibit the activity of tyrosine hydroxylase and tryptophan hydroxylase, the monoamine deficiency in the Qdpr-deficient mice were thought to be caused by hyperphenylalaninemia, not but a deficiency of BH4. The present study suggested that the monoaminergic neurons in the brain have ability to synthesize monoamine-neurotransmitters without any help of Qdpr.		3O1-02 Sex difference in hippocampal synapses and hormones Hojo Yasushi^1,2,3，Kato Asami^2,3，Chung Bon-chu³，Murakoshi Takayuki¹，Kawato Suguru^2,3 ¹Dept. of Biochem., Saitama Med. Univ.，²Grad. Sch. of Arts and Sci., Univ. of Tokyo，³JST, Japanese-Taiwanese Cooperative Programme Sex difference in the brain is a very attractive problem. For example, the hypothalamus, the brain region responsible for reproductive behavior, exhibits a clear sex difference in the size of nerve nucleus and the number of neurons. In contrast, the hippocampus, a center for learning and memory, does not have sex difference at the anatomical level including the volume and the number of neurons. Nevertheless, the significant sex difference in the performance of hippocampus-dependent task such as spatial memory using Morris water maze or radial arm maze task exists. We hypothesized that the sex difference in the hippocampal structure exists at more subtle level, that is, synaptic level. A novel software, Spiso-3D, which we developed, allowed us to reveal the sex difference in the density of spines（post synaptic region）and the fluctuation of spines in female with a period of 4 days（estrous cycle）. What generates the sex difference in the hippocampal synapses? So far, sex difference in the hippocampus has been attributed to the level of sex hormones in the blood. We revealed, however, that the hippocampal neurons themselves synthesized sex hormones including estradiol（E2）, testosterone（T）, dihydrotestosterone（DHT）and progesterone（PROG）in both sexes. The levels of sex hormones in hippocampus were higher than that in plasma, suggesting that hippocampal sex hormones had more impact to hippocampal functions than circulating ones did. The levels of sex hormones exhibited a clear sex difference, and in female, especially, fluctuated across estrous cycle. Surprisingly, no sex difference in the mRNA level and the localization pattern of steroidogenic enzymes and receptors was observed in the hippocampus. The estrous cycle-dependent fluctuation of the spine density in female rat hippocampus had a good correlation with the cyclic fluctuation of hippocampal levels of E2 and PROG. This clear sex difference in hormonal profile in hippocampus may generate the sex difference in the hippocampal structure at more subtle level, that is, synaptic level, resulting in the sex difference in the performance of hippocampus-dependent task.

3O1-03 Reduced axonal localization of a Caps2 splice variant impairs axonal release of BDNF and causes autistic-like behavior in mice Sadakata Tetsushi¹，Furuichi Teiichi² ¹Advanced Scientific Research Leaders Development Unit, Gunma University，²Department of Applied Biological Science, Tokyo University of Science Ca2+-dependent activator protein for secretion 2（CAPS2 or CADPS2）potently promotes the release of brain-derived neurotrophic factor（BDNF）. A rare splicing form of CAPS2 with deletion of exon3（dex3）was identified to be overrepresented in some patients with autism. Here, we generated Caps2-dex3 mice and verified a severe impairment in axonal Caps2-dex3 localization, contributing to a reduction in BDNF release from axons. In addition, circuit connectivity, measured by spine and interneuron density, was diminished globally. The collective effect of reduced axonal BDNF release during development was a striking and selective repertoire of deficits in social- and anxiety-related behaviors. Together, these findings represent the first mouse model of a molecular mechanism linking BDNF-mediated coordination of brain development to autism-related behaviors and patient genotype.		3O1-04 Rational design of a novel high-affinity, ultrafast, red calcium indicator R-CaMP2 Inoue Masatoshi^1,5，Takeuchi Atsuya²，Horigane Shin-ichiro¹，Ohkura Masamichi³，Gengyo-Ando Keiko³，Fujii Hajime¹，Kamijo Satoshi^1,5，Takemoto-Kimura Sayaka^1,4，Kano Masanobu²，Nakai Junichi³，Kitamura Kazuo^2,4，Bito Haruhiko^1,5 ¹Dept. of Neurochemistry, Grad. Sch. of Med.,. Univ. of Tokyo，²Dept. of Neurophysiol., Grad. Sch. of Med.,. Univ. of Tokyo，³Brain Science Institute, Saitama Univ.，⁴PRESTO-JST，⁵CREST-JST Fluorescent Ca²⁺ reporters are widely used as readouts of neuronal activities. Here, we designed R-CaMP2, a novel high-affinity red genetically encoded calcium indicator（GECI）with a Kd for Ca²⁺＜70 nM, and with a Hill coefficient near 1. Use of the calmodulin-binding sequence of CaMKK-α/β in lieu of a M13 sequence resulted in three fold faster kinetics than R-CaMP1.07 in rise and decay time of Ca²⁺ transients. These features allowed to resolve single action potential（AP）and fast AP trains up to near 20-40 Hz in acute cortical slices. In vivo imaging of the barrel cortex layer 2/3 neurons revealed reliable recording of single APs in R-CaMP2-expressing neurons, while synaptic Ca²⁺ transients were robustly detected in individual dendritic spines with similar efficacy as previously reported ultrasensitive green GECIs. R-CaMP2 exhibits a linear relationship between AP trains and fluorescence dynamics in vivo. Combining green and red GECIs, we successfully achieved dual-color monitoring of neuronal activities of distinct cell types, in the mouse cortex and in free-moving C. elegans. Together, R/G-CaMP imaging using R-CaMP2 provides a powerful means to interrogate orthogonal and hierarchical active ensembles, thus significantly enhancing our current capacity for functional mapping of neuronal circuits in vivo.